肿瘤相关巨噬细胞对人结肠癌SW620细胞生物学功能的影响

摘要

目的 研究肿瘤相关巨噬细胞(TAM)对人结肠癌SW620细胞生物学功能的影响.方法 采用白细胞介素(IL)- 4体外诱导人M2型巨噬细胞,Western blot检测其标志分子CD68、巨噬细胞甘露糖受体(MMR)和诱导型一氧化氮合酶(iNOS)的表达情况.Transwell非接触式共培养TAM和SW620细胞,酶联免疫吸附法(ELISA)检测TAM分泌的细胞因子IL-10、IL-12、IL-23和转化生长因子-β(TGF-β)水平;活性凝胶电泳迁移分析法(EMSA)检测SW620细胞中核因子-κB(NF-κB)活性;四氮甲唑蓝法(XTT)和荧光激活细胞分选术(FACS)双标记法分别检测培养后SW620细胞的增殖和凋亡情况.结果 IL- 4诱导的人M2型巨噬细胞可表达CD68和MMR,不表达iNOS.SW620与M2型巨噬细胞共培养24、48h后,培养上清液中M2型巨噬细胞分泌的IL-10和TGF-β水平较培养前明显升高(P均＜0.01),IL-12和IL-23水平与培养前差异无统计学意义(P均＞0.05).与M2型巨噬细胞共培养24、48h后,SW620的NF-κB DNA结合活性较培养前分别降低了72%和75%(P均＜0.01),细胞增殖活性分别下降了48%和59%(P均＜0.01);凋亡率分别为6.37%和7.68%,明显高于对照组的0.37%(P均＜0.01).结论 TAM可能通过抑制SW620细胞的NF-κB活性,抑制SW620细胞增殖,促进SW620细胞凋亡.%Objective To study the influence of tumor-associated macrophages (TAMs) on the biological function of SW620 cell. Methods Macrophage was induced into M2-type macrophage form with interleukin (IL)-4. CD68, macrophage mannose receptor (MMR), and inducible nitric oxide synthase (iNOS) were analyzed with Western blot. SW620 was co-cultured with TAMs in the Transwell. Cytokines including IL-10, IL-12, IL-23, and tramsforming growth factor-β (TGF-β) were detected with enzyme-linked immunosorbent assay (ELISA). The activity of nuclear factor-κB (NF-κB) in SW620 was analyzed with electrophoretic mobility shift assay (EMSA). The proliferation and apoptosis of SW620 cells after co-cultured with TAM were determined with tetrazolium four nitrogen (XTT) assay and fluorescence activated cell sorting ( FACS), respectively. Results IL-4 induced M2 type macrophage expressed CD68 and MMR instead of iNOS. After cocultured with SW620 for 24 hours and 48 hours, M2 type macrophage secreted higher levels of IL-10 and TGF-β than the pre-culture level ( P ＜ 0. 01 ), although IL-12 and IL-23 showed no significant differences ( P ＞ 0.05 ).The activity of NF-κB in SW620 decreased by 72％ and 75％ after 24 hours and 48 hours compared with the pre-culture level, respectively (both P ＜ 0.01 ). The activity of proliferation decreased by 48％ and 59％ and the apoptotic rates increased by 6.37 ％ and 7.68 ％ and 0.37 ％ after 24 hours and 48 hours ( all P ＜ 0.01 ) compared with the pre-culture levels. Conclusion TAM may inhibit the proliferation and promote the apoptosis of SW620 by suppressing the activity of NF-κB.